Chaperone activity and structure of monomeric polypeptide binding domains of GroEL

Abstract

The chaperonin GroEL is a large complex composed of 14 identical 57-kDa subunits that requires ATP and GroES for some of its activities. We find that a monomeric polypeptide corresponding to residues 191 to 345 has the activity of the tetradecamer both in facilitating the refolding of rhodanese and cyclophilin A in the absence of ATP and in catalyzing the unfolding of native barnase. Its crystal structure, solved at 2.5 Å resolution, shows a well-ordered domain with the same fold as in intact GroEL. We have thus isolated the active site of the complex allosteric molecular chaperone, which functions as a "minichaperone." This has mechanistic implications: the presence of a central cavity in the GroEL complex is not essential for those representative activities in vitro, and neither are the allosteric properties. The function of the allosteric behavior on the binding of GroES and ATP must be to regulate the affinity of the protein for its various substrates in vivo, where the cavity may also be required for special functions.

Accelerated folding in the weak hydrophobic environment of a chaperonin cavity: Creation of an alternate fast folding pathway

“…In addition, it has been observed that folding of hen lysozyme can be promoted by GroEL alone, in the absence of GroES and ATP (20). Similar results have been demonstrated for barnase (21) and for rhodanese (22).…”

Mycobacterium tuberculosis GroEL Homologues Unusually Exist as Lower Oligomers and Retain the Ability to Suppress Aggregation of Substrate Proteins

“…For example, functionally active monomeric minichaperones containing part of the polypeptide binding domain of GroEL are known to be effective in vitro and in vivo. 32,33 Also monomeric Cpn60 from Thermus thermophilus has been shown to possess the ability to suppress aggregation and promote protein folding with no requirement for ATP or GroES. 34 Thus, M. tuberculosis Cpn60s in a similar fashion might have evolved to function in lower oligomeric state with no ATP requirement.…”

ERp57 Is a Multifunctional Thiol-Disulfide Oxidoreductase

“…In this study, all ERp57 residue numbers refer to the human sequence after cleavage of the predicted signal peptide to generate the mature N terminus beginning with the sequence Ser 1 -Asp 2 -Val 3 . Gene products encoding the a and a domains were amplified by PCR and cloned into a pRSET A-derived E. coli expression vector (35). Protein expression from this vector results in the production of a fusion protein with an N-terminal 17-amino acid affinity tag containing a hexahistidine sequence.…”